1. Field of the Invention
The present invention relates to a method of selectively removing a protein from a biological sample using chemicals.
2. Description of the Related Art
Efficient extraction of DNA from cells is required in many applications and is essential in molecular diagnosis, in particular, the identification and quantification of pathogenic bacteria. Molecular diagnosis is generally performed by DNA amplification after DNA extraction. Types of DNA amplification include a polymerase chain reaction (PCR), a ligase chain reaction, a stranded-displacement amplification, a nucleic acid-based amplification, a repair chain reaction, a helicase chain reaction, a QB replicase amplification, and a ligation activated transcription.
The production of high purity double-strand plasmid DNAs, single-strand phage DNAs, chromosomal DNAs, and agarose gel-purified DNA fragments is very important in molecular biology. Ideal methods of purifying DNAs should be simple, be performed rapidly and include little additional manipulation of samples. The DNAs obtained using such methods are ready for direct transformation, restriction enzyme analysis, ligation, or sequencing. Such methods are commonly used in automated production of DNA samples, which is favored in research and diagnosis labs. Generally, the preparation of plasmid DNAs from crude alcohol precipitates is laborious. Plasmid DNAs are often produced using a CsCl gradient, gel filtration, ion exchange chromatography, RNAase, proteinase K, and repeated alcohol precipitation. These methods require considerable downstream sample preparation to remove CsCl and other salts, EtBr, and alcohol, etc. Further, small negatively charged cellular components can be precipitated together with DNAs. Thus, the DNAs may be contaminated to an undesirable degree.
In general, protein largely occupies a large part of a cell composition. Accordingly, in order to purify nucleic acids from biological samples, protein that largely forms a cell should be efficiently removed. It is known that trichloroacetic acid (TCA) can precipitate protein by three chloro groups contained therein (J. Prot. Chem. 1997, 16(4): 291-297.) However, precipitating agents that precipitate protein or nucleating agents that nucleate protein also precipitate nucleic acids, in addition to protein. For example, it is known that genome DNAs are precipitated by TCA (J. Biol. Chem. 1945, 161:293-303.) Accordingly, there is a need to develop a method capable of removing only protein while not removing nucleic acids from a biological sample containing protein. Inventors of the present invention researched a method of removing protein, based on conventional techniques, and found that when a protein nucleating agent is added to a sample together with a specific compound, such as betaine, and the resultant mixture is treated with a hydrophobic surface material, protein can be efficiently removed while a nucleic acid exists in the sample.